High-fidelity detection of DNA combining the CRISPR/Cas9 system and hairpin probe

• Published in: Biosensors & bioelectronics Vol. 184; p. 113212
• Main Authors: Wang, Meng, Han, Dongsheng, Zhang, Jiawei, Zhang, Rui, Li, Jinming
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 15.07.2021; Elsevier BV
• Subjects: Biosensing Techniques; biosensors; blood serum; Clustered Regularly Interspaced Short Palindromic Repeats; Clustered Regularly Interspaced Short Palindromic Repeats - genetics; CRISPR; CRISPR-Cas Systems; CRISPR-Cas Systems - genetics; detection limit; DNA; DNA - genetics; Gene Editing; genotyping; Hairpin probe; Isothermal amplification; Limit of Detection; Nucleic acid detection; nucleotide sequences; sequence analysis; SNVs genotyping; CRISPR; SNVs genotyping; Hairpin probe; Isothermal amplification; Nucleic acid detection
• DOI: 10.1016/j.bios.2021.113212
• ISSN: 0956-5663; 1873-4235

Methods that enable specific and sensitive detection of DNA are greatly required for high-fidelity sequence measurement and single-nucleotide variations (SNVs) genotyping. The CRISPR (clustered regularly interspaced short palindromic repeats)/Cas systems have provided revolutionary tools for detecting nucleic acids. However, most of the current CRISPR/Cas-based DNA biosensing platforms suffer from inherent off-target effects of Cas proteins and require pre-amplification processes, which compromise the analytical fidelity. In this work, a CRISPR/Cas9-triggered hairpin probe-mediated biosensing method (namely CHP) was used to directly read the original DNA sequences, while effectively neutralizing the off-target effect and achieving high sensitivity. This technique can quantify DNA targets with a limit of detection (LOD) at the attomole level and identify SNVs with allelic fractions as low as 0.01%~0.1%. Moreover, we show that the CHP system is applicable in detecting mutations in serum samples without DNA isolation steps. Collectively, the CHP system is a sensitive and high-fidelity platform, which promises a great potential for providing robust tool for DNA sequence analysis and SNVs genotyping. •The CHP system is a high-fidelity DNA biosensor combining CRISPR/Cas9 and hairpin probe.•The CHP system can neutralize the off-target effect of CRISPR/Cas9.•The CHP system can quantify DNA at attomole level and identify SNVs at low allelic fraction.•The CHP system can detect mutations in serum samples without DNA isolation steps.